Brassinosteroids regulate petal spur length in Aquilegia by controlling cell elongation.

BACKGROUND AND AIMS: Aquilegia produce elongated, three-dimensional petal spurs that fill with nectar to attract pollinators. Previous studies have shown that the diversity of spur length across the Aquilegia genus is a key innovation that is tightly linked with its recent and rapid diversification into new ranges, and that evolution of increased spur lengths is achieved via anisotropic cell elongation. Previous work identified a brassinosteroid response transcription factor as being enriched in the early developing spur cup. Brassinosteroids are known to be important for cell elongation, suggesting that brassinosteroid-mediated response may be an important regulator of spur elongation and potentially a driver of spur length diversity in Aquilegia. In this study, we investigated the role of brassinosteroids in the development of the Aquilegia coerulea petal spur. METHODS: We exogenously applied the biologically active brassinosteroid brassinolide to developing petal spurs to investigate spur growth under high hormone conditions. We used virus-induced gene silencing and gene expression experiments to understand the function of brassinosteroid-related transcription factors in A. coerulea petal spurs. KEY RESULTS: We identified a total of three Aquilegia homologues of the BES1/BZR1 protein family and found that these genes are ubiquitously expressed in all floral tissues during development, yet, consistent with the previous RNAseq study, we found that two of these paralogues are enriched in early developing petals. Exogenously applied brassinosteroid increased petal spur length due to increased anisotropic cell elongation as well as cell division. We found that targeting of the AqBEH genes with virus-induced gene silencing resulted in shortened petals, a phenotype caused in part by a loss of cell anisotropy. CONCLUSIONS: Collectively, our results support a role for brassinosteroids in anisotropic cell expansion in Aquilegia petal spurs and highlight the brassinosteroid pathway as a potential player in the diversification of petal spur length in Aquilegia.

A role for the Auxin Response Factors ARF6 and ARF8 homologs in petal spur elongation and nectary maturation in Aquilegia.

The petal spur of the basal eudicot Aquilegia is a key innovation associated with the adaptive radiation of the genus. Previous studies have shown that diversification of Aquilegia spur length can be predominantly attributed to variation in cell elongation. However, the genetic pathways that control the development of petal spurs are still being investigated. Here, we focus on a pair of closely related homologs of the AUXIN RESPONSE FACTOR family, AqARF6 and AqARF8, to explore their roles in Aquileiga coerulea petal spur development. Expression analyses of the two genes show that they are broadly expressed in vegetative and floral organs, but have relatively higher expression in petal spurs, particularly at later stages. Knockdown of the two AqARF6 and AqARF8 transcripts using virus-induced gene silencing resulted in largely petal-specific defects, including a significant reduction in spur length due to a decrease in cell elongation. These spurs also exhibited an absence of nectar production, which was correlated with downregulation of STYLISH homologs that have previously been shown to control nectary development. This study provides the first evidence of ARF6/8 homolog-mediated petal development outside the core eudicots. The genes appear to be specifically required for cell elongation and nectary maturation in the Aquilegia petal spur.

Homologs of LEAFY and UNUSUAL FLORAL ORGANS Promote the Transition From Inflorescence to Floral Meristem Identity in the Cymose Aquilegia coerulea.

Homologs of the transcription factor LEAFY (LFY) and the F-box family member UNUSUAL FLORAL ORGANS (UFO) have been found to promote floral meristem identity across diverse dicot model systems. The lower eudicot model Aquilegia produces cymose inflorescences that are independently evolved from the well-studied cymose models Petunia and tomato. We have previously characterized the expression pattern of the Aquilegia homolog AqLFY but in the current study, we add expression data on the two UFO homologs, AqUFO1 and 2, and conduct virus-induced gene silencing of all the loci. Down-regulation of AqLFY or AqUFO1 and 2 does not eliminate floral meristem identity but, instead, causes the transition from inflorescence to floral identity to become gradual rather than discrete. Inflorescences in down-regulated plants generate several nodes of bract/sepal chimeras and, once floral development does commence, flowers initiate several whorls of sepals before finally producing the wildtype floral whorls. In addition, silencing of AqUFO1/2 appears to specifically impact petal identity and/or the initiation of petal and stamen whorls. In general, however, there is no evidence for an essential role of AqLFY or AqUFO1/2 in transcriptional activation of the B or C gene homologs. These findings highlight differences between deeply divergent dicot lineages in the functional conservation of the floral meristem identity program.

Breaking the mold: understanding the evolution and development of lateral organs in diverse plant models.

The formation of complex three-dimensional shape differs significantly between plants and animals due to the presence of the cell wall in the former, which prevents all cell migration. Instead, in lateral plant organs such as leaves or petals, shape is controlled by a series of developmental phases in which the organ acquires polarity, cells undergo proliferation, and, lastly, cells expand to their final shape and size. Although these processes were first described based on mutagenesis approaches in major model systems like Arabidopsis thaliana, further insight into their complexity is best provided by studies of natural variation in organ shape in alternative model systems that sample a broader range of plant form. Weaving together work from both forward and evolutionary genetics, this review focuses on how modification in polarity establishment, cell proliferation and cell expansion drives modifications in the fundamental lateral organ developmental program to create diversity in shape.